Novobiocin sensitivity and chromatin conformation in wild-type and temperature-sensitive mouse L cells

Colwill, R.; Sheinin, Rose

doi:10.1139/o82-024

Cited by 6 publications

(4 citation statements)

References 7 publications

(13 reference statements)

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“…In all cases the DNA of cells, released to interact with concentrations of EtdBr of 1-20 /ug/mL, yielded the expected sedimentation pattern [cf. Mattern & Painter (1979) and Colwill & Sheinin (1982)]. This is seen in panel a of Figure 5 for the wild-type cells.…”

Section: Resultsmentioning

confidence: 71%

“…The EtdBr titration profiles for the chromatin-bound DNA of wild-type BalB/C-3T3 cells growing at 34 or 38.5 °C, and of ts 2 cells incubated at the pt or the npt, are reminiscent of those obtained with other wild-type (Brazell & Cook, 1975;Colwill & Sheinin, 1982;Mattern & Painter, 1979) and dnats mammalian cells (Colwill & Sheinin, 1982). They provide evidence for a supercoiled conformation for the nuclear DNA in BalB/C-3T3 and ts 2 mouse fibroblasts.…”

Section: Discussionmentioning

confidence: 70%

See 1 more Smart Citation

DNA synthesis in Balb/C-3T3 ts 2 cells is restricted by a temperature-sensitive function of late G1 phase

Sheinin¹,

Mirjah²,

Dubsky³

et al. 1986

Biochemistry

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ts 2 BalB/C-3T3 mouse fibroblasts are cdc mutants, which arrest late in G1, at or near the G1/S traverse, upon full expression of the heat-sensitive lesion. The kinetics of temperature inhibition of DNA synthesis in logarithmically growing cultures reveal three stages of heat inactivation. During the first generation time equivalent, normal semiconservative, semidiscontinuous replication proceeds but is reduced as cells exit and do not reenter S phase. During a second such period, a minimal rate of normal DNA synthesis is maintained. Thereafter, as the cells move into a third aborted cell division cycle, the rate of DNA synthesis increases. However, all semiconservative synthesis is then replaced by DNA repair replication. Temperature inactivation of the ts 2 protein results in shutdown of nuclear DNA synthesis. In contrast, normal replication of mitochondrial DNA proceeds at control rate throughout the first stage of temperature inactivation. Synthesis of this organellar genome is quantitatively reduced as the cells move into the second phase of heat inhibition. Titration of chromatin-bound DNA with ethidium bromide revealed that wild-type cells exhibit a changing DNA topology as the temperature is raised. Temperature-inactivated ts 2 cells behave as though their DNA has been topologically frozen in the configuration of control cells at or near entry into S phase.

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Section: Resultsmentioning

confidence: 71%

Section: Discussionmentioning

confidence: 70%

DNA synthesis in Balb/C-3T3 ts 2 cells is restricted by a temperature-sensitive function of late G1 phase

Sheinin¹,

Mirjah²,

Dubsky³

et al. 1986

Biochemistry

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“…Despite the burgeoning interest in mammalian cell cycle mutants (Table 2) few of the lesions responsible are biochemically defined. Exceptions to this are seen for ts A159, a topoisomerase II mutant (Colwill & Sheinin, 1982, and for ts 85 (Mita et al, 1980;. Surprisingly this latter mutant which arrests on G2 is defective in the ubiquitin pathway.…”

Section: Cell Cycle Mutantsmentioning

confidence: 91%

“…Cell cycle mutations in several prokaryotes have been (1977), Moser & Meiss (1975 Floros et al (1978), Rossini & Baserga (1978), Jonak & Baserga (1979), Rossini et al (1979) Talavera & Basilico (1977 Nishimoto & Basilico (1978) Wang (1974), Wang & Yin (1976) Wang (1976 Chen & Wang (1977) Chen & Wang (1977) Wissinger & Wang (1978 Liskay , Liskay (1974), Farber & Liskay (1974) Thompson et al (1970, 1971), Sheinin (1976a,b Sheinin et al (1978), Colwill & Sheinin (1982 L tscl Late S or G2 Setterfield et al (1978), Sheinin & Gutman (1977), Thompson et al (1971) described: these provide fundamental insights into theory (Goodwin, 1980) and the participating of the ion mechanisms of chromosome replication, growth and cell currents observed by Jaffe (1981). Further information on division (e.g.…”

Section: Cell Cycle Mutantsmentioning

confidence: 99%

Biochemistry of the cell cycle

Lloyd¹

1987

Biochemical Journal

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DNA polymerase‐primase complex in wild‐type and ts A1S9 mouse L‐cells, temperature‐sensitive for dna replication during cell cycle progression

Munsch

Sheinin

1990

Journal Cellular Physiology

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ts A1S9 mutant cells, derived from wild type WT-4 mouse L-cells, are temperature-sensitive (ts) for DNA synthesis and cell division. We try to determine the cause of the arrest of DNA replication in ts A1S9 cells at the nonpermissive temperature by comparing the modifications induced by the shift of temperature on the activity and the synthesis of DNA polymerase-alpha and DNA primase as a function of time. Forty-seven hours after temperature upshift DNA polymerase-alpha activity of ts A1S9 cells was inhibited by 90% while primase activity was barely detectable. By contrast, the activities of both enzymes increased to a plateau level in WT-4 cultured at either temperature and in ts A1S9 cells grown at the low permissive temperature. Study of the synthesis of DNA polymerase-alpha primase and of the structure of the enzyme complex during cell cycle progression was approached by immunoprecipitation of [35S]-labelled cells, with a specific monoclonal antibody directed against DNA polymerase-alpha. We have found that, irrespective of temperature of cultivation of WT-4 or ts A1S9 cells, this antibody precipitated polypeptides of 220, 186, 150, 110, 68-70, 60, and 48 kDa from cell extracts. With ts A1S9 cells cultivated at 38.5 degrees C for 48 hr the polypeptides of 220 and 186 kDa, associated with alpha-polymerase activity, were considerably more abundant than in the control cells, with a concomitant decline in the polypeptides of 60 and 48 kDa, implicated in primase activity. Thus the inhibition of DNA polymerase-alpha cannot be due to a decreased synthesis of the 186 kDa subunit but to its temperature inactivation. Consistent with a recent asymmetric dimeric model where polymerase-alpha complex and polymerase delta complex synthesize co-ordinately at the replication fork lagging and leading DNA strands, the observed alterations of polymerase-alpha and primase content explain the inhibition of DNA synthesis and the cell cycle arrest of the ts A1S9 cells at the nonpermissive temperature.

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Novobiocin sensitivity and chromatin conformation in wild-type and temperature-sensitive mouse L cells

Cited by 6 publications

References 7 publications

DNA synthesis in Balb/C-3T3 ts 2 cells is restricted by a temperature-sensitive function of late G1 phase

DNA synthesis in Balb/C-3T3 ts 2 cells is restricted by a temperature-sensitive function of late G1 phase

Biochemistry of the cell cycle

DNA polymerase‐primase complex in wild‐type and ts A1S9 mouse L‐cells, temperature‐sensitive for dna replication during cell cycle progression

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